Negative growth regulatory role of PTPN11/SHP2 in breast epithelial tumorigenesis.

Abstract

Drosophila model system has played a pivotal role in the identification of genes involved at all stages of tumorigenesis. An RNAi mediated screen of the Drosophila genome conducted in our laboratory identified putative negative regulators of growth in Epidermal Growth Factor Receptor (EGFR) and Yorkie (Yki) driven tumors. In this study, human orthologues of the RNAi mediated screen were subjected to detailed network analyses. Enrichment of the protein-protein interaction network among these putative growth regulators (in humans) identified major signaling pathways like the Hippo pathway and the mitogen-activated protein kinase (MAPK) signaling pathway to be involved in driving both EGFR and the Yes-associated protein 1 (YAP1) (a mammalian ortholog of Yki) mediated tumorigenesis. One of the identified negative growth regulators in the Yki screen but not a positive in the EGFR screen was corkscrew (Csw), a non-receptor protein tyrosine phosphatase. Tyrosine-protein phosphatase type 11 (PTPN11) encoding the Src Homology Phosphatase 2 (SHP2) protein is one of the closest human orthologs of Csw and it is reported to be a bonafide oncogene in EGFR driven neoplasia. However, our clinical metadata analysis showed PTPN11/SHP2 functions as a putative tumor suppressor in breast adenocarcinoma (BRCA). Our analysis of dataset availed by the Molecular Taxonomy of Breast cancer International Consortium (METABRIC), 2012 showed PTPN11/SHP2 copy number loss in Luminal A subtype of breast cancer patient correlated to poor 4 years disease-specific survival (DSS) and late-stage cancer at diagnosis. Furthermore, our analysis of The Cancer Genome Atlas (TCGA) BRCA level 4 protein data showed low expression levels of active phospho SHP2-Y542 associated with larger tumor size and more lymph node positivity in Luminal A subtype of patients at diagnosis. We experimentally investigated the possible role of PTPN11/SHP2 as a tumor suppressor in breast cancer. Knockdown of PTPN11/SHP2 in MCF10A, a non-transformed breast epithelial cell line, showed increased migration and cell shape changes to mesenchymal morphology, although there was no change in the rate of cell proliferation. Besides, the treatment of PTPN11/SHP2 knockdown cells with epirubicin (chemotherapeutic drug) showed better survival and reduced apoptosis. However, we did not observe any change in the levels of Bcl associated X protein (BAX), a global pro-apoptotic molecule. PTPN11/SHP2 may interact with other apoptotic effectors, which needs to be further investigated. 2 In the context of YAP1 driven neoplasia, we analyzed the METABRIC dataset. Cohorts that had high levels of YAP1 and copy number loss of PTPN11/SHP2 correlated to grade 3 tumors at diagnosis and poor 4 years DSS. We examined this relationship in YAP1 overexpressing MCF10A cells. Knockdown of PTPN11/SHP2 in the background of over-expressed YAP1 did not change the proliferation rate, which was already at higher levels when wild type YAP1 alone was over-expressed. In summary, we conclude, the tumor suppressor role of PTPN11/SHP2 in breast tumorigenesis may use a pathway independent of YAP1. Molecules like PTPN11/SHP2, YAP1, and P53 among many others which show dual specificity in tumorigenesis in the same tissue depending on the upstream signaling cues present challenges in the field of targeted drug therapy. This study puts forth the importance of understanding the contexts and behavior of such dual molecules depending on upstream signaling cues to allow effective cancer treatment.